In spite of recent medical progress, cancer continues to be one of the most common and deadly diseases. Elucidation of biochemical pathways involved in development and progression of various cancers is important to identify potential anti-cancer treatments as well as to develop agents effective to regulate such pathways in other aspects of health and disease.
Akt kinase family members, Akt1/PKBα, Akt2/PKBβ or Akt3/PKBγ, play important roles in development of a large variety of cancers including melanoma (Robertson, G. P., Cancer Metastasis Rev., 2005, 24:273-85; Stahl, J. M. et al., Cancer Res., 2004, 64:7002-10.)
A particular cancer, melanoma, is the most deadly form of skin cancer due to its high metastatic potential. Akt3 and downstream PRAS40 are part of a key signaling cascade activated in ˜70% of melanomas. Akt3 functions to reduce cellular apoptosis in early melanomas, thereby promoting development of this disease.
Inducible Nitric Oxide Synthase (iNOS) deregulation is associated with cancers, including melanoma, and is correlated with poor survival. iNOS is a calcium independent, cytokine-inducible enzyme involved in production of a bioactive, pleiotropic regulatory and signaling molecule from arginine called nitric oxide (Rao, C. V., Mutat. Res., 2004; 555:107-19.) Among other cancers, recent studies have implicated iNOS as a potential therapeutic target in melanoma (Ekmekcioglu, S. et al., Clin. Cancer Res., 2000, 6:4768-75; Ekmekcioglu, S. et al., Int J Cancer 2006; 119:861-6; Massi, D. et al., J. Pathol., 2001; 194:194-200; Salvucci, O. et al., Cancer Res., 2001, 61:318-26.)
Among the three isoforms of NOS, calcium independent iNOS produces high levels of nitric oxide promoting development of a malignant phenotype (Rao, C. V., Mutat. Res., 2004; 555:107-19; Ekmekcioglu, S. et al., Clin. Cancer Res., 2000, 6:4768-75; Ekmekcioglu, S. et al., Int J Cancer 2006; 119:861-6; Salvucci, O. et al., Cancer Res., 2001, 61:318-26.) Elevated iNOS expression/activity is also correlated with poor survival rates of melanoma patients (Ekmekcioglu, S. et al., Clin. Cancer Res., 2000, 6:4768-75; Ekmekcioglu, S. et al., Int J Cancer 2006; 119:861-6)
A number of small molecule iNOS inhibitors have been identified and tested in vitro and in viva (Rao, C. V., Mutat. Res., 2004, 555:107-19; Misko, T. P. et al., Eur J. Pharmacol., 1993; 233:119-25; Zheng, M. et al., Cancer Chemother Pharmacol 2007, 60:625-33; Garvey, E. P. et al., J. Biol. Chem., 1994, 269:26669-76.) In vitro studies using iNOS inhibitors, s-methylisothiourea and aminoguanidine, have demonstrated inhibition of nitric oxide production and induction of apoptosis mediated by caspase-1/3, and PARP cleavage (Salvucci, O. et al., Cancer Res., 2001, 61:318-26.)
PBIT is an iNOS selective inhibitor that is effective preventing colon and esophageal cancer in rats following dietary administration. However, due to low potency, poor cell permeability and associated systemic toxicity its utility in clinical settings is limited. (Garvey, E. P. et al., J. Biol. Chem., 1994, 269:26669-76; Rao, C. V., Mutat. Res., 2004, 555:107-19; Chen, T. et al., Cancer Res 2004; 64:3714-7.)
Constitutive activation of the MAP (Ras/Raf/Mek/Erk) kinase pathway through Ras mutations in 10-15% and B-Raf mutation in ˜60% of melanomas is a second important signaling cascade deregulated in cancers, including melanomas (Madhunapantula, S. V. et al., Cancer Res., 2008, 68:5-8.) Activating B-Raf mutations are acquired, somatic, post-zygotic events and are not inherited in families (Lang J. et al., Hum. Mutat., 2003, 21:327-30.) Among different B-Raf mutations, a single-base missense substitution (T to A at nucleotide 1799) that changes valine to glutamic acid at codon 600 (V600E) in exon 15 is prevalent in ˜90% of melanoma tumors with mutation of B-Raf (Davies, H. et al., Nature, 2002, 417:949-54). V600EB-Raf protein leads to kinase activity 10.7 times higher than occurs in normal cells and causes hyperactivity of the MAP kinase pathway (Davies, H. et al. Nature, 2002, 417:949-54).
Treatment of cells with PBIT to inhibit iNOS increases MAP kinase pathway activity leading to higher levels of active phosphorylated Erk1/2 and increased expression of downstream iNOS (Ellerhorst, J. A. et al., Oncogene, 2006, 25:3956-62; Tunctan B, et al., Pharmacol. Res., 2007, 56:56-64). Abnormally high activation of the MAP kinase pathway can inhibit cellular growth in a wide variety of normal and cancer cells by promoting cellular senescence, including melanomas (Michaloglou, C. et al., Nature, 2005, 436:720-4; and Dhomen, N. et al., Cancer Cell, 2009, 2009, 15, 294-303.). Recent evidence suggests that constitutively active V600EB-Raf initially promotes nevi development during melanoma tumor progression that results in high, intense activation of the MAP kinase pathway is inhibitory and that Akt3 activity is required to phosphorylate B-Raf in order to reduce its and the MAP kinase pathway activity to levels promoting rather than inhibiting proliferation (Cheung, M., et al., Cancer Res., 2008, 68:3429-3439).
Compositions and methods are required to inhibit abnormal cell survival and proliferation. In particular, compositions and methods are required to modulate signaling molecules Akt, iNOS and/or MAP kinase, and reduce survival and proliferation of abnormal cells such as cancer cells.